1. Field of the Invention
This invention relates to a device for use in automating the detection of target nucleic acid sequences in biological-containing samples. The device described herein is for use with an automated process, including a fluid-delivery system and a thermal reaction chamber, as is described in co-pending U.S. patent application Ser. No. 07/227,348. The disclosure of the co-pending application is hereby incorporated herein by reference.
2. Description of the Related Art
Devices for receiving biological specimens for diagnostic purposes are varied and adapted to the methods of detection. The devices may take the form of tubes for liquid specimens, flat surfaces such as glass slides suitable for microscopy, microtiter dishes, Petri dishes and cubes containing growth medium, or filters made of various materials to which cell and viral components will adhere
These specimen samples are then treated in such a way as to indicate either the presence or absence, or quantity, of a specific biological entity Test reagents may either be preapplied to the device or added in series after the specimen is present. Test results are read manually by a technical person or automatically with instrumentation specifically designed for that assay. In some instances the specimen is diluted with a diluent, or an aliquot of the specimen is removed from the original collecting device and transferred to another vessel at some point in the assay. In some cases physical and chemical means are used to amplify the signal of the assay for greater sensitivity Some assays require extraction or separation to isolate a specific component from other parts.
In DNA-based diagnostics the sequence specificity of base-pairing or enzymatic or other types of cleavage is exploited. The linear sequence of nucleotides in double-stranded DNA molecules forms the basis of replication of the genetic code. Hybridization is the binding of two single-stranded DNA strands whose base-pairing sequences are complementary. Temperature and salt concentration affect the stringency of these base-pairing matches A change from high stringency to low stringency can cause the same DNA probe to be either exquisitely specific to detect a particular target or less specific and detect a group of related targets.
In some instances the sizes of DNA fragments, produced by restriction endonuclease digestion or by amplification of a target sequences between primer pairs, are used to make a DNA-print for individual identification or aid in diagnosis of a genetic disease, cancer or infectious disease. For example, electrophoresis may be used to size-fractionate different-sized nucleic acids which have been specifically cleaved or whose native length puts them in a distinguishable size-length class. In the electrophoresis method, a current is applied to DNA loaded at the cathodal end of a gel matrix, which causes the DNA to migrate towards the anodal end of the matrix. The electrophoretic mobility of DNA is dependent on fragment size and is fairly independent of base composition or sequence. Resolution of one size class from another is better than 0.5% of fragment size (Sealy P. G. and E. M. Southern. 1982. Gel electrophoresis of DNA, p. 39-76. In D. Rickwood and B. D. Hames (EDS.), Gel Electrophoresis of Nucleic Acids. IRL Press, London). This reference and all other publications or patents cited herein are hereby incorporated by reference.
Electrophoresis methods thus require a vessel to hold the matrix material and the biological specimens to be subjected to electrophoresis. Such vessels may mold the gel matrix during its formation and may hold it during processing.
Diffusion of reagents is faster where the ratio of the matrix surface area to matrix volume is greatest as in thin, flat matrices. Likewise, electrophoresis of macromolecules requires less voltage and is faster in ultra-thin matrices or tiny (glass) capillaries. In these aqueous matrices, the vessel is necessary to prevent evaporation and to add strength in handling. Existing vessels that enclose matrices impede rapid diffusion of reagents and molecular probes. Once the existing vessels are taken apart in processing, they cannot be put back together to continue automated processing.
Accordingly, the invention aims to provide a vessel for the individual specimens to be contained.
Yet another object of the invention is to mold matrix material which is to contain the specimen.
A further object of the invention is to carry the specimen in transport from the point of collection to the processing point.
A further object of the invention is to provide support of the specimen, embedding it in a matrix for automated processing.
A further object of the invention is to provide a convenient way to make the particles containing target nucleic acids of a specimen in a matrix available for optimal signal detection.
A further object of the invention is to allow for saturating specimens quickly with a series of solutions or for drying them automatically.
A further object of the invention is to concentrate specimen nucleic acids, or amplified products thereof, for detection of their presence.
A further object of the invention is provide a barrier to evaporation of solutions during processing.
A further object of the invention is a mechanism to change its configuration during processing of the specimen to adapt to processing conditions.
A further objective of the invention is to provide support for reading the test results.
A still further object of the invention is to permanently store the nucleic acids present in the specimen for possible retesting and serve as a permanent record of the test, if an archival record is desired.
Other objects and advantages of the invention will be more fully apparent from the ensuing disclosure and appended claims.